Use single cell RNA sequencing to study the molecular siganature of primary human Megakaryocytic-erythroid progenitor
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE112692 | GEO | 2018/11/02
REPOSITORIES: GEO
Similar Datasets
Project description:The differentiation of primary human megakaryocyte-erythroid progenitors (MEP) into megakaryocytic (Mk) or erythroid (E) progenitors is critical for maintaining the blood system, yet the mechanisms that regulate this fate specification remain unclear. Here, we analyzed RNA-seq data and found that RUNX1 is a key regulator of differential gene expression in MEP fate specification. Through viral transduction of primary MEP with RUNX1, we demonstrated that overexpression of RUNX1 promotes Mk over E specification, whereas pan-RUNX inhibition promotes E fate specification over Mk. We found that although the levels of total RUNX1 do not differ between MkP and ErP, MkP have higher levels of RUNX1 phosphorylated serine residues, and that serine/threonine phosphorylation of RUNX1 dramatically increases its effectiveness. We used human erythroleukemia (HEL) cell lines with expression of HA-tagged WT RUNX1, phosphomimetic (RUNX1-4D) and non-phosphorylatable (RUNX1-4A) mutants to model their effects on MEP. Although the chromatin association was not informative, the three forms of RUNX1 caused differential expression of 2,625 genes. Approximately 40% of these differentially expressed genes (DEGs) showed an increase with both RUNX1-WT and RUNX1-4D while another 40% showed a decrease with both RUNX1-WT and RUNX1-4D. We compared these DEGs with DEGs from primary human MEP, Mk progenitors (MkP), and E progenitors (ErP), and found that the genes upregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes upregulated in MkP vs. MEP, whereas genes downregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes downregulated in MkP vs. MEP. These results suggest that phosphorylation of RUNX1 enhances RUNX1’s function to activate or repress transcription of target genes that are up/downregulated during MEP fate specification.
2024-09-24 | GSE232878 | GEO
Project description:The differentiation of primary human megakaryocyte-erythroid progenitors (MEP) into megakaryocytic (Mk) or erythroid (E) progenitors is critical for maintaining the blood system, yet the mechanisms that regulate this fate specification remain unclear. Here, we analyzed RNA-seq data and found that RUNX1 is a key regulator of differential gene expression in MEP fate specification. Through viral transduction of primary MEP with RUNX1, we demonstrated that overexpression of RUNX1 promotes Mk over E specification, whereas pan-RUNX inhibition promotes E fate specification over Mk. We found that although the levels of total RUNX1 do not differ between MkP and ErP, MkP have higher levels of RUNX1 phosphorylated serine residues, and that serine/threonine phosphorylation of RUNX1 dramatically increases its effectiveness. We used human erythroleukemia (HEL) cell lines with expression of HA-tagged WT RUNX1, phosphomimetic (RUNX1-4D) and non-phosphorylatable (RUNX1-4A) mutants to model their effects on MEP. Although the chromatin association was not informative, the three forms of RUNX1 caused differential expression of 2,625 genes. Approximately 40% of these differentially expressed genes (DEGs) showed an increase with both RUNX1-WT and RUNX1-4D while another 40% showed a decrease with both RUNX1-WT and RUNX1-4D. We compared these DEGs with DEGs from primary human MEP, Mk progenitors (MkP), and E progenitors (ErP), and found that the genes upregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes upregulated in MkP vs. MEP, whereas genes downregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes downregulated in MkP vs. MEP. These results suggest that phosphorylation of RUNX1 enhances RUNX1’s function to activate or repress transcription of target genes that are up/downregulated during MEP fate specification.
2024-09-24 | GSE232877 | GEO
Project description:The differentiation of primary human megakaryocyte-erythroid progenitors (MEP) into megakaryocytic (Mk) or erythroid (E) progenitors is critical for maintaining the blood system, yet the mechanisms that regulate this fate specification remain unclear. Here, we analyzed RNA-seq data and found that RUNX1 is a key regulator of differential gene expression in MEP fate specification. Through viral transduction of primary MEP with RUNX1, we demonstrated that overexpression of RUNX1 promotes Mk over E specification, whereas pan-RUNX inhibition promotes E fate specification over Mk. We found that although the levels of total RUNX1 do not differ between MkP and ErP, MkP have higher levels of RUNX1 phosphorylated serine residues, and that serine/threonine phosphorylation of RUNX1 dramatically increases its effectiveness. We used human erythroleukemia (HEL) cell lines with expression of HA-tagged WT RUNX1, phosphomimetic (RUNX1-4D) and non-phosphorylatable (RUNX1-4A) mutants to model their effects on MEP. Although the chromatin association was not informative, the three forms of RUNX1 caused differential expression of 2,625 genes. Approximately 40% of these differentially expressed genes (DEGs) showed an increase with both RUNX1-WT and RUNX1-4D while another 40% showed a decrease with both RUNX1-WT and RUNX1-4D. We compared these DEGs with DEGs from primary human MEP, Mk progenitors (MkP), and E progenitors (ErP), and found that the genes upregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes upregulated in MkP vs. MEP, whereas genes downregulated by WT and RUNX1-4D in HEL cells overlapped significantly with the genes downregulated in MkP vs. MEP. These results suggest that phosphorylation of RUNX1 enhances RUNX1’s function to activate or repress transcription of target genes that are up/downregulated during MEP fate specification.
2024-09-24 | GSE232879 | GEO
Project description:The mechanism underlying thrombocytosis in patients with iron deficiency anemia remains unknown. We present findings that support the hypothesis that low iron biases the commitment of Megakaryocytic-Erythroid Progenitors (MEP) toward the megakaryocytic (Mk) lineage in mice. In MEP of Transmembrane serine protease 6 knockout (Tmprss6-/-) mice, which exhibit iron deficiency microcytic anemia concomitant with thrombocytosis, we observed a megakaryocytic (Mk) bias, decreased labile iron, and decreased proliferation relative to wild-type (WT) MEP. Bone marrow transplantation assays suggest that the systemic iron deficiency of the Tmprss6-/- recipients contributes to the MEP lineage commitment bias. Genes involved in metabolic, VEGF, and ERK pathways were enriched among those differentially expressed between WT and Tmprss6-/- MEP. Corroborating our findings from the murine model of chronic iron deficiency anemia, primary human MEP also exhibited decreased proliferation and Mk-biased commitment when their iron sensing pathways were disrupted by knockdown of Transferrin Receptor 2. These data are consistent with a model in which low iron in the marrow environment affects MEP metabolism, attenuates ERK signaling, slows proliferation, and biases MEP toward Mk lineage commitment. Keywords: megakaryocytic-erythroid progenitor (MEP), iron deficiency, Tmprss6
2023-03-17 | GSE227563 | GEO
Project description:Using single-cell RNA sequencing (scRNAseq), we investigated ETV6-variant (P214L and F417LTer4) carriers who are affected by constitutional thrombopenia. We demonstrate that ETV6-variant carriers presented transcriptomic aberrant populations of megakaryocytes (MK) and mega-erythroid progenitors (MEP) with affected pathways. We confirmed this bioinformatic analyzes by functional assays. We also demonstrate a bypass of common myeloid progenitors (CMP) during physiologic differentiation from hematopoietic stem/progenitors cells (HSPC) to MK and we identified a new population of MK-primed CMP in our culture model. We raise this analysis as a new exploratory modus operandi for constitutional diseases.
2023-06-15 | GSE206089 | GEO
Project description:The common myelo-erythroid progenitor (CMP) was subdivided based on expression of CD27. To understand the relationship between the various CD27 fractions of the CMP and downstream progenitors (granulocyte-monocyte progenitor, GMP, and megakaryocyte-erythroid progenitor, MEP), genome-wide transcriptional analysis was used.
2016-10-31 | GSE33937 | GEO
Project description:Background: Recent advances in single-cell techniques have provided the opportunity to finely dissect cellular heterogeneity within populations previously defined by “bulk” assays and to uncover rare cell types. In human hematopoiesis, megakaryocytes and erythroid cells differentiate from a shared precursor, the megakaryocyte-erythroid progenitor (MEP), which remains poorly defined.Results: To clarify the cellular pathway in erythro-megakaryocyte differentiation, we correlated the surface immunophenotype, transcriptional profile and differentiation potential of individual MEP cells. Highly purified, single MEP cells (n=681) were analyzed using index fluorescence-activated cell sorting with parallel targeted transcriptional profiling of the same cells performed using a specifically designed panel of 87 genes. Differentiation potential was tested in novel, single-cell differentiation assays. Our results demonstrated that immunophenotypic MEP in fact comprise three distinct subpopulations: (1) “Pre-MEP”, enriched for erythroid/megakaryocyte progenitors but with residual myeloid differentiation capacity (2) “E-MEP”, strongly biased towards erythroid differentiation, and (3) “MK-MEP”, a previously undescribed, rare population of cells that are bipotent but primarily generate megakaryocytic progeny. Therefore, conventionally-defined MEP are in fact a mixed population: a minority give rise to mixed-lineage colonies while the majority of cells are transcriptionally-primed to generate exclusively single-lineage output. Conclusions: Our study clarifies the cellular hierarchy in human megakaryocyte/erythroid lineage commitment and highlights the importance of using a combination of single-cell approaches to dissect cellular heterogeneity and identify rare cell types within a population. We present a novel immunophenotyping strategy that enables the prospective identification of specific intermediate progenitor populations in erythro-megakaryopoiesis, allowing for in-depth study of disorders including inherited cytopenias, myeloproliferative disorders and erythromegakaryocytic leukemias.
2016-04-02 | GSE79331 | GEO
Project description:MEIS1 is a transcription factor expressed in hematopoietic stem and progenitor cells (HSPC) and in mature megakaryocytes. In contrast to its role in leukemogenesis, the role of MEIS1 in normal hematopoiesis is largely unknown. We show that MEIS1 can direct human hematopoietic progenitors towards a megakaryocyte-erythroid progenitor (MEP) fate. Ectopoic expression of MEIS1 in CD34+ cells resulted in increased erythroid differentiation at the expense of granulocyte and monocyte (GM) differentiation. MEIS1 overexpression not only skewed differentiation of CMPs towards the erythroid lineage but also reprogrammed GM progenitors towards erythrocyte differentiation. Expression profiling was used to determine the transcriptional changes induced by MEIS1 that lead to the oberved phenotype. A transcriptional program enriched for erythrocytic and megakaryocytic genes was detected.
2014-11-12 | GSE53263 | GEO
Project description:Background: Recent advances in single-cell techniques have provided the opportunity to finely dissect cellular heterogeneity within populations previously defined by âbulkâ assays and to uncover rare cell types. In human hematopoiesis, megakaryocytes and erythroid cells differentiate from a shared precursor, the megakaryocyte-erythroid progenitor (MEP), which remains poorly defined.Results: To clarify the cellular pathway in erythro-megakaryocyte differentiation, we correlated the surface immunophenotype, transcriptional profile and differentiation potential of individual MEP cells. Highly purified, single MEP cells (n=681) were analyzed using index fluorescence-activated cell sorting with parallel targeted transcriptional profiling of the same cells performed using a specifically designed panel of 87 genes. Differentiation potential was tested in novel, single-cell differentiation assays. Our results demonstrated that immunophenotypic MEP in fact comprise three distinct subpopulations: (1) âPre-MEPâ, enriched for erythroid/megakaryocyte progenitors but with residual myeloid differentiation capacity (2) âE-MEPâ, strongly biased towards erythroid differentiation, and (3) âMK-MEPâ, a previously undescribed, rare population of cells that are bipotent but primarily generate megakaryocytic progeny. Therefore, conventionally-defined MEP are in fact a mixed population: a minority give rise to mixed-lineage colonies while the majority of cells are transcriptionally-primed to generate exclusively single-lineage output. Conclusions: Our study clarifies the cellular hierarchy in human megakaryocyte/erythroid lineage commitment and highlights the importance of using a combination of single-cell approaches to dissect cellular heterogeneity and identify rare cell types within a population. We present a novel immunophenotyping strategy that enables the prospective identification of specific intermediate progenitor populations in erythro-megakaryopoiesis, allowing for in-depth study of disorders including inherited cytopenias, myeloproliferative disorders and erythromegakaryocytic leukemias. Multiplex RT-PCR gene expression profiling of 807 human megakaryocyte-erythroid progenitor cells (MEP) isolated from three healthy donors by apheresis following G-CSF treatment. Cells were excluded if more than 70 assays did not result in amplification or displayed Ct higer than 13 for B2M or higher than 15 for GAPDH. Furthermore cells with a mean non-dropout Ct value greater than 20 were removed. This resulted in a dataset of 681 cells, which were subsequently normalised to the mean of B2M and GAPDH expression.
2016-04-02 | E-GEOD-79331 | biostudies-arrayexpress
Project description:Megakaryopoiesis is a complex process that involves major cellular changes and relies on controlled coordination of cellular proliferation and differentiation. These mechanisms are orchestrated in part by transcriptional regulators. The key hematopoietic transcription factor SCL/TAL1 is required for specification of the megakaryocytic lineage from hematopoietic progenitors. Here, we report that it also critically controls terminal megakaryocyte maturation. In vivo depletion of SCL specifically in the megakaryocytic lineage affects all key attributes of megakaryocyte progenitors (MkPs), namely proliferation, ploidisation, cytoplasmic maturation, as well as platelet production. Genome-wide expression analysis reveals increased expression of the cell cycle regulator p21 in Scl-deleted MkPs. Importantly, P21 knockdown-mediated rescue assays in Scl-deleted MkPs show full restoration of cell cycle progression and partial rescue of the nuclear and cytoplasmic maturation defects. Therefore, SCL-mediated transcriptional control of p21 is critical for maturation of MkPs. Our study provides a mechanistic link between one of the major hematopoietic transcriptional regulators, cell cycle progression and megakaryocytic differentiation. Total RNA extracted from control TGPF4-CRE;SCL wt/wt sorted MK progenitors was compared to total RNA extracted from TGPF4-CRE;SCL fl/fl MK progenitors cells where Scl was completely excised
2011-07-11 | E-GEOD-24969 | biostudies-arrayexpress